Detection, quantification, isolation and purification of target biomaterials, such as viruses and biomacromolecules (including constituents or products of living cells, for example, proteins, carbohydrates, lipids, and nucleic acids) have long been objectives of investigators. Detection and quantification are important diagnostically, for example, as indicators of various physiological conditions such as diseases. Isolation and purification of biomacromolecules, such as monoclonal antibodies produced from cell cultures or fermentation processes, are important for therapeutic and in biomedical research. Biomacromolecules such as enzymes have been isolated, purified, and then utilized for the production of sweeteners, antibiotics, and a variety of organic compounds such as ethanol, acetic acid, lysine, aspartic acid, and biologically useful products such as antibodies and steroids.
Chromatographic separation and purification operations can be performed on biological product mixtures, based on the interchange of a solute between a moving phase, which can be a gas or liquid, and a stationary phase. Separation of various solutes of the solution mixture is accomplished because of varying binding interactions of each solute with the stationary phase; stronger binding interactions generally result in longer retention times when subjected to the dissociation or displacement effects of a mobile phase compared to solutes which interact less strongly and, in this fashion, separation and purification can be effected.
Most current capture or purification chromatography is done via conventional column techniques. These techniques have severe bottlenecking issues in downstream purification, as the throughput using chromatography is low. Attempts to alleviate these issues include increasing the diameter of the chromatography column, but this in turn creates challenges due to difficulties of packing the columns effectively and reproducibly. Larger column diameters also increase the occurrence of problematic channeling. Also, in a conventional chromatographic column, the absorption operation is shut down when a breakthrough of the desired product above a specific level is detected. This causes the dynamic or effective capacity of the adsorption media to be significantly less than the overall or static capacity. Furthermore, a selective Protein A column must be protected from unwanted contamination that may plug the column making it difficult to clean or possibly ruining the column for reuse. This reduction in effectiveness has severe economic consequences, given the high cost of some chromatographic resins.
Polymeric resins are widely used for the separation and purification of various target compounds. For example, polymeric resins can be used to purify or separate a target compound based on the presence of an ionic group, based on the size of the target compound, based on a hydrophobic interaction, based on an affinity interaction, or based on the formation of a covalent bond.
There is a need in the art for functionalized membranes that overcome limitations in diffusion and binding, and that may be operated at high throughput and at lower pressure drops. There is a need in the art for polymeric substrates having enhanced affinity for selective removal of biocontaminates, such host cell proteins, cell debris, DNA fragments, viruses and cell debris from biological feed-streams in the production of monoclonal antibodies.
There is further need in the art for functionalized membranes that have low levels of total organic extractables. USP Tests are used to determine the biological reactivity of elastomerics, plastics, and other polymeric materials. These tests are detailed in the general chapters Biological Reactivity Tests, In Vitro and Biological Reactivity Tests, In Vivo in the US Pharmacopeia. The Biological Reactivity Tests, In Vitro are designed to determine the biological reactivity of mammalian cell culture following contact with polymeric materials with direct or indirect patient contact or of specific extracts prepared from the materials under test.
According to the injection and implantation testing requirements specified under Biological Reactivity Tests, In Vivo, plastics and polymers are graded on a scale of Class I to Class VI. To grade a plastic or polymer, extracts of the test material are generated in various media and are injected systematically and intracutaneously into rabbits or mice to evaluate their biocompatibility. An additional level of implantation testing may be performed. Plastics not requiring implantation testing are graded Class I, II, III, or V and those plastics requiring implantation testing are graded Class IV or VI. The USP procedure for each test outlines the “pass” criteria so that it can be said that a particular product sample meets the requirements of the test. One significant concern is the amount of organic compounds that may be extracted from such polymers, described as “total organic carbon” or TOC.